This grant request is focused on how 11-cis retina is biosynthesized in the retina/pigmented epithelium of vertebrate animals. The question of how this process occurs is formulated in two separate but coupled ways. The first objective is to determine the mechanism of 11-cis retinal biosynthesis. Of great importance here is to determine whether the isomerization of an all trans retinoid to an 11-cis retinoid is enzyme-mediated or not. The second objective is to determine where the energy comes from for the biosynthesis of the 11-cis retinoids. In a dark adapted eye, greater than 75% of the retinoids can be in the 11-cis form, whereas at equilibrium, only 0.1% is in the 11-cis form. The formation of equal quantities of 11-cis and all-trans retinoids requires the input of approximately 4 kcal/mol. The question of the mechanism of 11-cis retinoid formation will be approached by determining 1) the in vivo stereospecificity of the isomerization process, 2) which all-trans retinoid is isomerized, and 3) whether any evidence for the material presence of a retinoid isomerase can be obtained. If evidence for an enzyme is found, the enzyme will be purified and characterized. Whether an isomerase enzyme exists or not, energy source for the production of 11-cis retinoid must be found. Our ultimate goal here is to determine whether the 11-cis retinal(o1) binding protein can provide the energy or whether an ATP-dependent mechanism might be involved. The research alluded to here could be of importance in understanding the etiology of certain forms of night blindness and retinitis pigmentosa.